The principal aim of this investigation is to understand the regulation of expression of the genes and enzymes involved in the vitamin K biosynthetic pathway. Vitamin K is an important fat soluble vitamin that is required for the well being of humans and animals. Vitamin K responsive hypoprothrombia is an important clinical problem under a number of conditions. The bacterial flora of the human intestine contribute from 50-100% of the human vitamin K requirement. Vitamin K is derived from the "shikimate pathway" and it is formed from isochorismate and 2-ketoglutarate. The intermediates that have been identified are 2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC), o-succinylbenzoate (OSB), a CoA ester of OSB (OSB-CoA), 1,4- dihydroxy-2-naphthoate (DHNA), and desmethylmenaquinone (DMK). The menD, C, E, and B genes located at 48.5 min of the chromosome have been cloned and sequenced. The sequencing of the menA gene at 89 min is in progress and we will identify and clone the gene for the conversion of DMK---> MK. To overcome the difficulty of working with small quantities of enzymes and intermediates, we will use operon and protein fusions to study regulation and we will introduce the genes into overexpression vectors to overproduce, purify and study the enzymes and intermediates of the pathway.